Choline acetyltransferase (commonly abbreviated asChAT, but sometimesCAT) is atransferaseenzyme responsible for the synthesis of theneurotransmitteracetylcholine. ChAT catalyzes the transfer of anacetyl group from thecoenzymeacetyl-CoA tocholine, yielding acetylcholine (ACh). ChAT is found in high concentration incholinergicneurons, both in thecentral nervous system (CNS) andperipheral nervous system (PNS). As with most nerve terminal proteins, ChAT is produced in the body of theneuron and is transported to thenerve terminal, where its concentration is highest. Presence of ChAT in a nerve cell classifies this cell as a "cholinergic" neuron. In humans, the choline acetyltransferase enzyme is encoded by theCHATgene.[5]
Choline acetyltransferase was first described byDavid Nachmansohn and A. L. Machado in 1943.[6] A German biochemist, Nachmansohn had been studying the process of nerve impulse conduction and utilization of energy-yielding chemical reactions in cells, expanding upon the works of Nobel laureatesOtto Warburg andOtto Meyerhof onfermentation,glycolysis, andmuscle contraction. Based on prior research showing that "acetylcholine's actions on structural proteins" were responsible for nerve impulses, Nachmansohn and Machado investigated the origin of acetylcholine.[7]
An enzyme has been extracted from brain and nervous tissue which forms acetylcholine. The formation occurs only in presence ofadenosinetriphosphate (ATP). The enzyme is called choline acetylase.
Theacetyltransferase mode of action was unknown at the time of this discovery, however Nachmansohn hypothesized the possibility of acetylphosphate or phosphorylcholine exchanging the phosphate (fromATP) for choline or acetate ion.[6] It was not until 1945 thatCoenzyme A (CoA) was discovered simultaneously and independently by three laboratories,[8][9][10] Nachmansohn's being one of these. Subsequently, acetyl-CoA, at the time called “active acetate,” was discovered in 1951.[11] The 3D structure of rat-derived ChAT was not solved until nearly 60 years later, in 2004.[12]
The 3D structure of ChAT has been solved by X-ray crystallographyPDB:2FY2. Choline is bound in the active site of ChAT by non-covalent interactions between the positively charged amine of choline and the hydroxyl group of Tyr552, in addition to ahydrogen bond between choline'shydroxyl group and ahistidine residue, His324.
The choline substrate fits into a pocket in the interior of ChAT, while acetyl-CoA fits into a pocket on the surface of the protein. The 3Dcrystal structure shows the acetyl group of acetyl-CoA abuts the choline binding pocket – minimizing the distance between acetyl-group donor and receiver.
Structure of choline acetyltransferase binding sites
Crystal structure of choline ion bound in choline acetyltransferase. Side chain residues of His324A and Tyr552A shown.PDB:2FY3
Stereoscopic depiction of choline and acetyl-CoA in ChAT active site.(PDB:2FY3,PDB:2FY5 - overlaid).
Stereoscopic depiction of choline and acetyl-CoA bound in ChAT active site - alternate angle. (PDB:2FY3,PDB:2FY5 - overlaid).
ChAT is very conserved across the animal genome. Among mammals, in particular, there is very high sequence similarity. Human and cat (Felis catus) ChAT, for example, have 89% sequence identity.Sequence identity withDrosophila is about 30%.[13]
There are two forms of ChAT: Soluble form and membrane-bound form.[14] The soluble form accounts for 80-90% of the total enzyme activity while the membrane-bound form is responsible for the rest of 10-20% activity.[15] However, there has long been a debate on how the latter form of ChAT is bound to the membrane.[16] The membrane-bound form of ChAT is associated with synaptic vesicles.[17]
There exist two isoforms of ChAT, both encoded by the same sequence. The common type ChAT (cChAT) is present in both the CNS and PNS. Peripheral type ChAT (pChAT) is preferentially expressed in the PNS in humans, and arises fromexon skipping (exons 6–9) duringpost-transcriptional modification. Therefore, theamino acid sequence is very similar; however, pChAT is missing parts of the sequence present in cChAT. The pChAT isoform was discovered in 2000 based on observations that brain-derived ChAT antibodies failed to stain peripheral cholinergic neurons as they do for those found in the brain. Thisgene splicing mechanism which leads to cChAT and pChAT differences has been observed in various species, including both vertebrate mammals and invertebrate mollusks, suggesting this mechanism leads to some yet-unidentified evolutionary advantage.
Cholinergic systems are implicated in numerous neurologic functions. Alteration in some cholinergic neurons may account for the disturbances ofAlzheimer disease. The protein encoded by this gene synthesizes the neurotransmitteracetylcholine. Acetylcholine acts at two classes of receptors in thecentral nervous system –muscarinic andnicotinic – which are each implicated in different physiological responses. The role of acetylcholine at the nicotinic receptor is still under investigation. It is likely implicated in the reward/reinforcement pathways, as indicated by the addictive nature ofnicotine, which also binds to the nicotinic receptor. The muscarinic action of acetylcholine in the CNS is implicated in learning and memory. The loss of cholinergic innervation in theneocortex has been associated with memory loss, as is evidenced in advanced cases of Alzheimer's disease. In theperipheral nervous system, cholinergic neurons are implicated in the control of visceral functions such as, but not limited to, cardiac muscle contraction and gastrointestinal tract function.
Mutants of ChAT have been isolated in several species, includingC. elegans,Drosophila, and humans. Most non-lethal mutants that have a non-wild type phenotype exhibit some activity, but significantly less than wild type.
InC. elegans, several mutations in ChAT have been traced to the cha-1 gene. All mutations result in a significant drop in ChAT activity. Percent activity loss can be greater than 98% in some cases. Phenotypic effects include slowed growth, decreased size, uncoordinated behavior, and lack of sensitivity towardcholinesterase inhibitors.[19] Isolatedtemperature-sensitive mutants inDrosophila have all been lethal. Prior to death, affected flies show a change in behavior, including uncontrolled movements and a change inelectroretinogram activity.[20]
The human gene responsible for encoding ChAT is CHAT. Mutations in CHAT have been linked tocongenital myasthenic syndrome, a disease which leads to general motor function deficiency and weakness. Further symptoms include fatalapnea. Out of ten isolated mutants, 1 has been shown to lack activity completely, 8 have been shown to have significantly decreased activity, and 1 has an unknown function.[21]
TheAlzheimer's disease (AD) involves difficulty in memory and cognition. The concentrations of acetylcholine and ChAT are remarkably reduced in the cerebral neocortex and hippocampus.[22] Although the cellular loss and dysfunction of the cholinergic neurones is considered a contributor to Alzheimer disease, it is generally not considered as a primary factor in the development of this disease. It is proposed that the aggregation and deposition of theBeta amyloid protein, interferes with the metabolism of neurones and further damages the cholinergic axons in the cortex and cholinergic neurones in the basal forebrain.[23]
Theamyotrophic lateral sclerosis (ALS) is one of the most common motor neuron diseases. A significant loss of ChAT immunoreactivity is found in ALS.[24] It is hypothesized that the cholinergic function is involved in an uncontrolled increase of intracellular calcium concentration whose reason still remains unclear.[25]
Neostigmine methylsulfate, an anticholinesterase agent, has been used to target ChAT. In particular, use of neostigmine methylsulfate has been shown to have positive effects against congenital myasthenic syndrome.[26]
Exposure toestradiol has been shown to increase ChAT in female rats.[27]
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